Human Molecular Genetics, 2015, Vol. 24, No. 19 5378–5387 doi: 10.1093/hmg/ddv261 Advance Access Publication Date: 10 July 2015 Original Article ORIGINAL ARTICLE Novel POC1A mutation in primordial dwarfism reveals Downloaded from https://academic.oup.com/hmg/article-abstract/24/19/5378/583656 by guest on 11 March 2020 new insights for centriole biogenesis Asuman Koparir1,†, Omer F. Karatas2,†, Betul Yuceturk3, Bayram Yuksel4, Ali O. Bayrak3, Omer F. Gerdan3, Mahmut S. Sagiroglu3, Alper Gezdirici5, Koray Kirimtay6, Ece Selcuk6, Arzu Karabay6, Chad J. Creighton7, Adnan Yuksel9 and Mustafa Ozen1,8,9,* 1Department of Medical Genetics, Istanbul University, Cerrahpasa Medical School, Istanbul, Turkey, 2Molecular Biology and Genetics Department, Erzurum Technical University, Erzurum, Turkey, 3Advanced Genomics and _ 4 Bioinformatics Research Center (IGBAM), BILGEM, TUBITAK, Kocaeli, Turkey, Genetic Engineering and Biotechnology Institute, TUBITAK Marmara Research Center, Kocaeli, Turkey, 5Department of Medical Genetics, Kanuni Sultan Suleyman Training and Research Hospital, 34303 Istanbul, Turkey, 6Department of Molecular Biology and Genetics, Istanbul Technical University, Istanbul, Turkey, 7Department of Medicine and Dan L Duncan Cancer Center, Division of Biostatistics and, 8Department of Pathology and Immunology, Baylor College of Medicine, Houston, TX 77030, USA and 9Department of Molecular Biology and Genetics, Biruni University, Topkapi, Istanbul, Turkey *To whom correspondence should be addressed. Tel: +90 2124448222; Fax: +90 2124164646; Email: [email protected] Abstract POC1A encodes a WD repeat protein localizing to centrioles and spindle poles and is associated with short stature, onychodysplasia, facial dysmorphism and hypotrichosis (SOFT) syndrome. These main features are related to the defect in cell proliferation of chondrocytes in growth plate. In the current study, we aimed at identifying the molecular basis of two patients with primordial dwarfism (PD) in a single family through utilization of whole-exome sequencing. A novel homozygous p.T120A missense mutation was detected in POC1A in both patients, a known causative gene of SOFT syndrome, and confirmed using Sanger sequencing. To test the pathogenicity of the detected mutation, primary fibroblast cultures obtained from the patients and a control individual were used. For evaluating the global gene expression profile of cells carrying p.T120A mutation in POC1A, we performed the gene expression array and compared their expression profiles to those of control fibroblast cells. The gene expression array analysis showed that 4800 transcript probes were significantly deregulated in cells with p.T120A mutation in comparison to the control. GO term association results showed that deregulated genes are mostly involved in the extracellular matrix and cytoskeleton. Furthermore, the p.T120A missense mutation in POC1A caused the formation of abnormal mitotic spindle structure, including supernumerary centrosomes, and changes in POC1A were accompanied by alterations in another centrosome-associated WD repeat protein p80-katanin. As a result, we identified a novel mutation in POC1A of patients with PD and showed that this mutation causes the formation of multiple numbers of centrioles and multipolar spindles with abnormal chromosome arrangement. † These authors contributed equally. Received: May 11, 2015. Revised and Accepted: July 4, 2015 © The Author 2015. Published by Oxford University Press. All rights reserved. For Permissions, please email: [email protected] 5378 Human Molecular Genetics, 2015, Vol. 24, No. 19 | 5379 Introduction In the current study, we utilized whole-exome sequencing (WES) in our PD patients to screen all disease-related and candi- Primordial dwarfism (PD) is a group of genetically heterogeneous date genes to find out the molecular etiology of the syndrome. disorders, and the term is used for describing prenatal onset A novel p.T120A homozygous missense mutation in exon 4 of growth failure, which persists postnatally. PD is divided into POC1A, which is a causative gene of SOFT syndrome, was de- two groups according to the head circumferences (HCs) of the tected. The variants were confirmed using Sanger sequencing, affected individuals: microcephalic and normocephalic. Micro- and functional analysis of the mutant allele was performed cephaly, associated with reduced brain size, is accompanied using the fibroblast cells cultured from skin biopsies of the pa- with most PD disorders including Seckel syndrome (MIM no. tients. In addition, we tested whether the mutation we identified 210600), microcephalic osteodysplastic PD (MOPD) type I (MIM affects the function of the centrosome during mitosis, and given no. 210710) and type II (MIM no. 210720) and Meier–Gorlin syn- the seemingly redundant and somewhat similar functions of drome (MIM no. 224690), which were generally related with intel- POC1 and p80-katanin, we aimed at investigating the changes lectual disability, except for MOPDII. There are other syndromes in p80-katanin protein in response to non-functional POC1A pro- Downloaded from https://academic.oup.com/hmg/article-abstract/24/19/5378/583656 by guest on 11 March 2020 such as Cornelia de Lange and Bloom’s syndromes, which are tein due to the deleterious mutation in POC1A. We also review characterized by prenatal onset growth retardation and micro- SOFT patient studies from the literature and describe additional cephaly; however, patients diagnosed with these syndromes also two Turkish siblings, one of whom had tall vertebral bodies and have other distinctive features. In contrast, a few PD disorders are precocious puberty, which might expand the phenotype. described with normal HC and usually normal intelligence includ- ing 3M syndrome (MIM no. 273750), Silver–Russell syndrome (MIM no. 180860) and Mulibrey nanism (MIM no. 253250). Short stature, Results onychodysplasia, facial dysmorphism and hypotrichosis (SOFT) Subjects syndrome (MIM no. 614813), which was described recently, is incompatible with this classification, as HC is normal in childhood The family has two children who are referred to us for short and getting smaller with advancing ages (1). stature by Endocrinology Department. The mother and father To date, more than 10 genes have been associated with micro- are first-degree cousins (Supplementary Material, Fig. S1). Both cephalic PD. Three of these genes are as follows: PCNT, CENPJ pregnancies were uncomplicated, with spontaneous vaginal (synonym with CPAP) and CEP152, which are related to centroso- deliveries at the term. The babies had low birth weight with mal functions (2–4). Although SOFT syndrome is not a microceph- roughly 1800 g and short length. alic PD, the only associated gene with this syndrome, POC1A, The first sibling of the family is 12-year-old female. At the age encodes POC1 centriolar protein A, which localizes to centrioles of 9, she was diagnosed with central precocious puberty and she and spindle poles (1). Centrosome integrity and duplication are was treated with Lucrin Depot, a gonadotropin-releasing hor- highly critical for cell cycle as microtubules use centrosomes as mone agonist. Cranial and pituitary magnetic resonance imaging anchoring points to extend from both poles in bipolar spindle in was normal. She had short stature, dolichocephaly, sparse hair, order to capture the chromosomes. Therefore, centrosome target- broad-wide forehead, sparse eyebrows and eyelashes, prominent ing is the most critical step for correct spindle formation, and given antihelix, large and prominent nose, full lips, large mouth, the established role of POC1A in centriole formation and mainten- high-arched palate, prognatism, brachydactyly, nail hypoplasia, ance, the gene is expected to have abnormalities in the mitotic prominent heels and pes planus (Fig. 1A–E). Skeletal surveys spindle in the presence of the deleterious mutations. showed slender long bones and tall vertebral bodies (Fig. 1J–N). Centrosome localization is the primary driving factor for micro- When she is referred to us at 9 years, her length, weight and HC tubule organization in both interphase and mitotic cells as centro- were 121 cm (−2 SD), 26.5 kg (25 centile) and 51.7 cm (25–50 somes serve as microtubule organization centers, and mutations centile), respectively. Her length for age and weight for age in centrosome-associated proteins underlie defects in microtubule percentiles were demonstrated in Supplementary Material, organization resulting abnormal spindle formation. WD40 micro- Figure S2A and B. tubule-associated proteins have long been known for their roles The second sibling is 8-year-old male. His physical examina- in centrosome targeting. In addition to POC1A, among many tions were similar with the first child having short stature, dolicho- other centrosome-related microtubule-associated proteins is kata- cephaly, broad-wide forehead, prominent antihelix, prominent nin (5). Katanin is a dimeric protein, which consists of p60 and nose, large mouth, high-arched palate, prognatism, brachydactyly, p80 subunits. P60/KATNA1 is a microtubule-severing enzyme nail hypoplasia, prominent heels and pes planus (Fig. 1AandF–I). that functions in the release of centrosome-born microtubules Radiographic images revealed slender long bones (Fig. 1O–T). Meta- and regulates microtubule length in both mitotic spindle and bolic screenings, echocardiography, abdominal ultrasonography, non-dividing/interphase cells. P80/KATNB1 is also a WD40 domain ophthalmic and otorhinolaryngological